TRIM5a is an antiviral host protein the mechanism of which is not yet understood. Rhesus macaque TRIM5a restricts HIV-1 but not SIV. Genetic and biochemical evidence suggest that TRIM5a binds directly to the viral capsid as the virus enters a target cell. Evidence was recently reported suggesting that the binding of TRIM5a to the incoming capsid causes a rapid uncoating of the virus, possibly resulting in a disruption to reverse transcription. The structure of TRIM5 suggests that it may act as an E3 ligase although a role for ubiquitination or capsid degradation has not been demonstrated. The proposed project seeks to derive understanding of the mechanism by which TRIM5a functions by identifying cellular proteins that are required for its antiviral function. Cellular proteins that are required for TRIM5a antiviral function will be identified by a genome-wide screen with an siRNA library that targets most of the expressed sequences in the human genome. The screen will detect siRNAs that interfere with rhesus TRIM5a function. As a secondary approach, cell proteins that form a complex with TRIM5a will be identified biochemically. The role of the proteins identified in the screen in mediating TRIM5a function will be studied by cell and molecular experiments. The specific aims of the project are the following: 1. Identify genes that are required for TRIM5a function by RNAi screening. 2. Identify proteins that physically associate with TRIM5a. 3. Determine the role of the gene products in TRIM5a antiviral function. Human cells contain a protein called TRIM5a that has the ability to prevent infection with HIV. TRIM5a is thought to attack the virus as it enters a cell by binding directly to it, although precisely how this works is not understood. The project will use a recently developed technology termed RNAi which allows researchers to selectively shut-off various functions of the cell, to discover the mechanism by which TRIM5a inhibits HIV infection. This understanding can be useful for the development of new drugs for AIDS. [unreadable] [unreadable] [unreadable]